Transfer ribonucleic acid was hydrolyzed by acid under conditions that released the purine residues. Examination of the hydrolysate by means of ion-exchange chromatography on a sulfonic acid resin revealed the presence of a A number of articles reports the presence of amino acids or small polypeptides, apart from the amino acids attached to the acceptor end of tRNA molecules, bound to nucleic acids. Ingram and Sullivan (1962) and Akashi et al. (1965), for example, have reported the presence of amino acids bound to RNA which cannot be removed through the use of extensive deproteinizing procedures. Balk et al. (1964) and Olenick and Hahn (1964) have reported the presence of amino acids in highly purified preparations of DNA isolated from a variety of sources. The nature of the amino acid-nucleic acid linkage, however, has not been elucidated. Bogdanov et al. (1962) have reported that tRNA contains amino acids attached to the phosphate residues. Harris and Wiseman (1962) have also reported the presence of small polypeptides attached to the

A ribonuclease from Ustilago sphaerogena (ATCC 12421) was purified 1100-fold with a yield of 32%. The isolated enzyme was free of contaminating phosphatases, ribonucleases, and deoxyribonucleases. The rate and specific-

The single-strand specific nuclease S1 from Aspergillus oryzae (EC 3.1.4.21) was purified 600-fold in 16% yield from dried mycelia. Determination of the isoelectric point of S1 nuclease as 4.3-4.4 allowed adjustment of chromatographic conditions such that the enzyme was isolated free of contaminating ribonucleases T1 and T2. S1 nuclease so purified was used for removal of single-stranded portions from the RNA of the Escherichia coli phage MS2, which has a helical content of about 65% in vitro. At 23 degrees, increasing amounts of enzyme converted the RNA to mononucleotides in about equimolar base ratios. No small intermediates of chain length 2-8 were found. At 0 degrees, MS2 RNA hydrolysis was slower and reached, in exhaustive digests, a plateau where 70% of the substrate RNA remained insoluble in 66% EtOH. With [32P]MS2 RNA, strip chart counting of 6% acrylamide-6 M urea electrophoresis patterns of such digests gave recoveries of 80-91% in the form of defined oligomer bands. On 2.5% acrylamide-0.5% agarose gels, the molecular weights of the major oligomers were found to range from 25,000 to 41,000. Similar to purified tRNAArg used as a control, these oligomers were not resistant to pancreatic RNase-RNase T1 hydrolysis at 37 degrees, and were not bound on hydroxylapatite at 50 degrees in 0.14 M sodium phosphate (pH 6.8). Melting of the oligomers gave complex profiles without a clear Tm and showed an increase in A260 of 35% at 93 degrees over that at 28 degrees. Upon formaldehyde denaturation of MS2 RNA prior to S1 nuclease hydrolysis, no resistant oligomers were found.

No abstract